Abstract
YerA41 is a Myoviridae bacteriophage that was originally isolated due its ability to infect Yersinia ruckeri bacteria, the causative agent of enteric redmouth disease of salmonid fish. Several attempts to determine its genomic DNA sequence using traditional and next generation sequencing technologies failed, indicating that the phage genome is modified in such a way that it is an unsuitable template for PCR amplification and for conventional sequencing. To determine the YerA41 genome sequence, we performed RNA-sequencing from phage-infected Y. ruckeri cells at different time points post-infection. The host-genome specific reads were subtracted and de novo assembly was performed on the remaining unaligned reads. This resulted in nine phage-specific scaffolds with a total length of 143 kb that shared only low level and scattered identity to known sequences deposited in DNA databases. Annotation of the sequences revealed 201 predicted genes, most of which found no homologs in the databases. Proteome studies identified altogether 63 phage particle-associated proteins. The RNA-sequencing data were used to characterize the transcriptional control of YerA41 and to investigate its impact on the bacterial gene expression. Overall, our results indicate that RNA-sequencing can be successfully used to obtain the genomic sequence of non-sequencable phages, providing simultaneous information about the phage–host interactions during the process of infection.
Highlights
YerA41 is a bacteriophage that was originally isolated due its ability to infect Yersinia ruckeri bacteria, the causative agent of enteric redmouth disease of salmonid fish [1]
The host range of YerA41 was studied on bacteria grown both at RT and at 37 ◦ C, as temperature is known to regulate surface structures in the genus
We show that a transcriptomic approach can be used to obtain the genomic sequence of infectious organisms that cannot be sequenced using traditional DNA based sequencing approaches as they may possess hypermodified deoxyribonucleotides
Summary
YerA41 is a bacteriophage that was originally isolated due its ability to infect Yersinia ruckeri bacteria, the causative agent of enteric redmouth disease of salmonid fish [1]. Bacteriophage YerA41 was first described in 1984 as a tailed icosahedral virus that lysed the vast majority of tested Y. ruckeri serovar I strains, and was believed to have a potential value for the diagnosis of redmouth disease. It was found that YerA41 has a relatively broad host range among Enterobacteriacae as it could infect some strains of other Yersinia species, Escherichia coli, Shigella flexneri, Enterobacter cloacae, Klebsiella and Erwinia spp. The YerA41 phage particles are large with heads of about 110 nm in diameter, 10 × 8 nm necks, 250 × 20 nm non-contracted sheaths and 70 nm long tail fibers [2]. Several attempts to sequence the YerA41 genome using generation sequencing (Illumina) approaches failed previously.
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